脱水淫羊藿素雌激素样效应调控Hedgehog信号通路影响易发肾虚型PMOP人群hUiPSCs骨向分化研究

Postmenopausal osteoporosis(PMOP) originate in estrogenic deficiency. Traditional Chinese Medcine considered that kidney determining the condition of the bone and kidney weakness lead to withered bone, so the therapeutic regimen for PMOP included reinforcing kidney herb epimedium. Anhydroicaritin(AHI), which sourced different origin of epimedium, had estrogenic effect and promoted mesenchymal stem cells(MSCs) osteogenic diferentiation. Hedgehog(Hh) signaling pathway regulates skeletal development and metabolism. Therefore, We hypothesized that estrogenic effect of AHI affect hUiPSCs osteogenic diferentiation through regulates Hh signaling pathway. This study is based on AHI and Hh signaling pathway, establishing susceptible PMOP (Kidney Weakness) hUiPSCs and induced MSCs as models, to investigate the regularities of AHI’s estrogenic effects induced PMOP hUiPSCs hedgehog-mediated osteogenic diferentiation, providing novel evidences for interpretation of kidney determining the condition of the bone and pharmacological mechanism of Chinese herb epimedium.

雌激素缺乏为绝经后骨质疏松症（PMOP）主要病因，中医认为“肾主骨”，“肾虚”则“骨萎”，治疗常用补肾中药淫羊藿，不同产地淫羊藿的共有成分为脱水淫羊藿素（AHI），具有雌激素效应，并促进间充质干细胞（MSCs）骨向分化。Hh信号调控骨组织发育与代谢，我们推测AHI雌激素效应通过调控Hh信号而影响其骨向分化。本研究从AHI与Hh信号切入，建立易发PMOP（肾虚型）人群hUiPSCs并诱导成MSCs为模型，研究雌激素效应调控Hh信号特点，阐明AHI干预hUiPSCs骨向分化调控Hh信号分子网络机制，为中医理论“肾主骨”及淫羊藿药理作用提供新依据。

我们成功建立了正常（NFe-UiPS）与肾虚型PMOP尿液诱导iPS（PMOP-hUiPSC）细胞系，在同时加经典骨向分化诱导剂的前提下，通过检测成骨分化相关指标，发现与正常人尿液来源NFe-UiPS相比，肾虚型PMOP-hUiPSC细胞具有较低的成骨分化能力；Western-blot检测PMOP-hUiPSC与NFe-UiPS胞内ERα与Hh信号相关蛋白表达水平，发现肾虚型PMOP-hUiPSC胞内ERα表达水平显著下调，而Hh信号通路被抑制；进一步我们明确了正常情况下， 0.1μmol/L、1μmol/L和10μmol/L这三个浓度AHI可显著促尿液诱导iPS增殖与骨向分化药理作用前提下，发现正常情况下，尿液诱导iPS骨向分化时，Hh信号激活，而最佳浓度10μmol/L的AHI可通过雌激素受体ERα进一步激活Hh信号而促进其骨向分化药理作用的具体分子机制。